Despite the tremendous knowledge related to the growth and transcription factors that regulate tooth development, there is still a lack of information related to the factors involved at later stages of tooth development, when the mineralized tissues are formed, and an even greater paucity of information as to the factors that regulate root development. It is the long-term objective of this study to understand the process of root development at the morphologic, cellular and molecular levels including the mechanisms underlying root dentin and cementum formation as well as root attachment to periodontal ligament in the healthy periodontium. In this application, we will focus on the role of Hertwig's Epithelial Root Sheath (HERS) cells in root development. Although the role of HERS cells in root formation is widely accepted, the precise function and fate of these cells remains controversial. Functions suggested for these cells have ranged from a pure structural entity (subdivide the dental ectomesenchymal tissues into dental papilla and dental follicle), to regulators of the timing of root development, to inducers for mesenchymal cells differentiation into cementoblasts, to being cementoblast cells precursors. In this competing renewal we will test the hypothesis that that root development is initiated by HERS cells through dynamic and reciprocal interactions with the surrounding mesenchyme resulting in budding morphogenesis and there is a unique gene associated with these tissues that is regulated by the NFI-C transcription factor. We propose to test this hypothesis by taking advantage of a newly created Nfi-c (Nuclear Factor I-C) knockout mice knock whose unique phenotype is that it lacks the formation of roots. In addition we will use established immortal cell lines (HERS, Dental Papillae Mesenchyme (DPM), dental follicle) to accomplish the following Specific Aims: To determine the initial events leading to root morphogenesis; To determine what kind of cell interactions are necessary for root development; To determine the molecules associated with root development; To determine the role of the molecules identified in root development. The use of a wide range of technology including in situ hybridization, tissue recombination, organ culture, cell culture, microarrays technology, etc. will provide the data required to accomplish our specific alms. The data generated in this study will provide much needed information related to root development, the role of HERS in this process and the regulatory factors involved in root formation. Although rare, individuals with root agenesis have been documented.